Escherichia coli is incapable of performing post-translational modifications; still, it has been the most common host system employed for the production of recombinant human therapeutic proteins. Among them, skin proteins including epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF), which do not require post-translational modifications such as glycosylation for biological activities and stability, have been expressed in E. coli using various tactics.
Probably due to its small size, EGF has been efficiently expressed, secreted and even excreted to the culture medium in E. coli. Excreted EGF was not only demonstrated to be bioactive, but more importantly, it was also shown to possess the correct primary (authentic) structure. Furthermore, EGF produced by excretion is free from endotoxin contamination as production of it through excretion does not require cell disruption. In addition, the low level of cytoplasmic proteins in the medium greatly facilitates the purification of EGF to high homogeneity. Importantly, the process is highly reproducible and easily scalable.
With respect to bFGF, despite its secretory nature, it has not been successfully expressed through either secretion or excretion in E. coli. Conventionally, expression of bFGF in E. coli has been confined to the cytoplasm using a fusion approach, in which retrieval of bFGF relies on cell lysis, proteolytic digestion of the fusion intermediate, and subsequent purification commonly accomplished by affinity chromatography. Intracellular expression of bFGF incurs high production costs as it requires either an expensive protease to cleave the fusion product or a labor-intensive protocol to regenerate bFGF when inclusion aggregates formed. Despite these manipulations, bFGF expression in E. coli has not been satisfactorily fulfilled as the resulting product was shown to be incorrectly processed or biologically inactive.
There had been attempts to use other recombinant systems to produce bFGF. One such attempt concerns the engineering of Bacillus subtilis to facilitate secretory production of bFGF. With the use of the NPR signal peptide of a B. amyloliquefaciens protease gene, which enabled accurate peptidase processing of the fusion precursor, together with a facile cell-wall destabilization protocol, bFGF possessing the authentic structure and full bioactivity was shown to be secreted into the culture medium in B. subtilis. 
The present invention provides an alternative and/or an improved methodology and system for producing bFGF and/or other useful polypeptides reliably and efficiently.